The present invention relates generally to the field of imaging, and more specifically, to imaging techniques in which an image is generated from the difference of two images made with light of different polarizations.
An image may be defined to be a mapping of the spatial distribution of some property of the specimen being imaged into another spatial distribution of the property or of a different property. For example, a typical black and white photographic negative of an object is a mapping of the spatial distribution of the intensity of light leaving each point on the object. The mapping process consists of setting the density of silver grains at each point in the negative in proportion to the intensity of light leaving the corresponding point on the specimen.
Imaging systems based on the mapping of a number of different physical properties are well known to the prior art. Such systems typically employ some form of electromagnetic radiation to probe the sample being imaged. For example, an x-ray image is a mapping of the ability of each point in the imaged part of the specimen to absorb electromagnetic radiation in the x-ray frequency range. A simple NMR image of a patient is a mapping of the hydrogen ion density at each point in a plane intersecting the patient's body.
Images of many biological specimens of interest are difficult to obtain because the structures one wishes to image do not significantly differ from the surrounding structures in their absorbance of electromagnetic radiation. For example, protein and DNA have similar densities and gross chemical compositions; hence it is difficult to distinguish one from the other using imaging based on the absorption of light.
One prior art solution to imaging biological specimens is to stain the specimen with a specific stain which binds to the structure of interest and which is easily imaged. For example specific stains have been developed which bind to specific proteins. These stains are typically constructed by combining a monoclonal antibody with a chromophore which is easily detected using light of a specific frequency. Although such staining systems provide greatly improved images in those cases in which they are applicable, they typically provide only information about the concentration of the particular protein to which the antibody binds. No information is provided about the orientation of the protein in question or its organization into larger structures unless the organization alters the density of the protein molecules significantly. Such staining systems also may produce artifacts resulting from the disruption of the specimen needed to introduce the staining agent or from the non-preferential binding of the stain to other structures in the specimen. In addition, a specific stain must be made for each imaging problem, which involves considerable time and expense.
Further, in a large number of imaging problems, one does not know in advance the best parameter to use to construct an image which distinguishes two specimens, one normal and one abnormal. If the two specimens differed in the concentration of some known compound, an imaging system based on a stain might be possible. However, in general, no such compound is known in advance and, hence, one is left to try a number of different imaging modalities in an attempt to find one that distinguishes the two specimens. This will be especially difficult if the two specimens differ mainly in the organization of some compound rather than in its concentration.
In principle, organizational differences may be detected using polarized light. A structure in the image composed of molecules which are bundled with a specific orientation will absorb light of one polarization in preference to light of another polarization. Hence an image formed by subtracting a first image taken with polarized light having one linear polarization from a second image taken with polarized light having a different linear polarization will often display such structures.
Similarly, man biological molecules of interest are chiral in nature. A chiral structure is one whose mirror image is different from the image of the structure. For example, helical molecules such as DNA are either left or right handed depending on the twist of the helix. The mirror image of a right handed helix is a left handed helix, not a right handed helix. Such molecules preferentially absorb circularly polarized light of one handedness. Hence an image formed from the difference of two images, one taken with right handed circularly polarized light and the other taken with left handed circularly polarized light will, in principle, display the concentration of such molecules.
Such differential image techniques also find application in the material sciences. For example, strains in transparent materials such as glass may be detected with polarized light. Here, specific staining techniques are of little value.
Unfortunately, the magnitude of the difference of the absorbance of circularly polarized light of different handedness for most biological specimens is quite small at wavelengths in the optical region of the light spectrum. Often, to measure this difference in the visible region of the spectrum, each of the images must be measured to an accuracy of the order of one part in ten thousand. In principle, one could increase this difference by using smaller wavelength light. The maximum difference in absorbance occurs at wavelengths which are of the same length as the diameter of the molecules or structures being imaged. Thus to obtain the maximum difference for chiral molecules, circularly polarized light in the x-ray region of the spectrum is needed. However, circularly polarized light sources in this region of the spectrum are not economically practical.
Prior art systems which attempt to exploit this type of differential imaging are often limited by the sensitivity of the detector used to form the two images which are subtracted. For example, if a standard television camera tube is used to record each of the images, differences which are less than a few percent of the light intensity of each image may not be detected. This limit is imposed by the small dynamic range of the television camera and by the electronic noise in the camera and the subsequent amplifiers. Hence it is difficult to apply this type of imaging technique to biological samples unless large concentrations of chiral molecules are present or unless large ordered structures are being imaged.
Broadly, it is an object of the present invention to provide an improved system for producing images which are the difference of two images made using light of different polarizations.
It is a further object of the present invention to provide an imaging system which can detect differences of less than one part in a thousand in the two images.
These and other objects of the present invention will become apparent from the following detailed description of the invention and the accompanying drawings.